We are studying a group of related parvoviruses which differ in host range as a result of a small number of differences in their capsid proteins. Canine parvovirus (CPV) emerged in 1978 as a variant of feline panleukopenia virus (FPV), and then underwent further host adaptation. FPV and CPV capsids bind the transferrin receptors (TfR) of their host cells, and while both can bind the feline TfR and use that for infection, CPV can also bind and use the canine TfR. The canine host range of CPV was due to a small number of differences in the capsid, and then the virus acquired further adaptive capsid changes. The TfRs bind the virus through the receptor apical domain, and CPV-specific binding involved a new glycosylation site. The TfR directs the virus into the correct endosomal pathway and prepares the capsid for cell infection. Public health significance: These studies address the fundamental aspects of how viruses use cellular processes to infect cells, and how evolution creates new host ranges and allows the emergence of new epidemic strains. The 2 aims address different aspects of virus-cell recognition leading to infection and controlling host ranges. Aim 1: Determine how the interactions between viral capsids and the feline and canine TfRs control cell infection and determine the viral host ranges. We would define the processes by which feline and canine TfRs and mutants of those receptors bind wild type and mutant viruses and mediate infection. Virus would be selected on mutant forms of the TfR and the adapted viruses examined for changes in sequence, and the structural changes in the capsids modeled. The flexibility of the capsids and the structural consequences of the TfR-capsid binding would be examined by assaying for protease sensitivity of the capsid, receptor, and of capsid-TfR complexes. Entry of viruses would be compared in various host cells, and the effects of varying the interactions on the uptake and endosomal transport of the capsids would be examined. The role of glycosylations of the canine TfR would be examined by changing that site and by testing receptors for virus binding and infection. Aim 2: To define how changes specifically in endosomal uptake and trafficking lead the virus into the correct pathway for infection and regulate viral host ranges. Here we would specifically examine the effects of receptor trafficking on infection and host range by examining the effects of mutant TfRs with altered TM or cytoplasmic sequences. Viruses lacking phospholipase A2 activity or VP1, or with changes around the fivefold axis, would be compared for their endosomal localization and release. We would seek to complement endosomal escape mutants in trans by adding adenovirus capsids, competent wild type parvovirus, or transferrin-PEI conjugates to the cells along with the parvoviruses.